The process of sealing of damaged axons was examined in isolated strips of
white matter from guinea pig spinal cord by recording the "compound membran
e potential," using a sucrose-gap technique, and by examining uptake of hor
seradish peroxidase (HRP). Following axonal transection, exponential recove
ry of membrane potential occurred with a time constant of 20 +/- 5 min, at
37 degrees C, and extracellular calcium activity ([Ca2+](o)) of 2 mM. Most
axons excluded HRP by 30 min following transection. The rate of sealing was
reduced by lowering calcium and was effectively blocked at [Ca2+](o) less
than or equal to 0.5 mM, under which condition most axons continued to take
up HRP for more than 1 h. Sealing at higher [Ca2+](o) was blocked by calpa
in inhibitors (calpeptin and calpain inhibitor-1) indicating a requirement
for type II (mM) calpain in the sealing process. Following compression inju
ry, the amplitude of the maximal compound action potential conducted throug
h the injury site was reduced. The extent of amplitude reduction was increa
sed when the tract was superfused with calcium-free Krebs' solution (Ca2+ r
eplaced by Mg2+). These results suggest that the fall in [Ca2+](o) seen fol
lowing injury in vivo is sufficient to prevent membrane sealing and may par
adoxically contribute to axonal dieback, retrograde cell death, and "second
ary" axonal disruption.